The cytoskeletal framework appears to have a role in cell shape, cell motility, anchorage-dependent growth, distribution of cell surface proteins, and the localization of organelles such as mitochondria and polyribosomes. The goal of this project is to identify and characterize proteins associated with the cytoskeletal framework of human lung cancer cells which differ from each other in morphology, adherence characteristics, surface membrane composition, and capacity to synthesize certain enzymes and polypeptide hormones. By two-dimensional gel electrophoresis of double-detergent insoluble cellular fractions, small-cell carcinoma lines were found to contain a different set of cytoskeleton-associated proteins from non-small lung carcinoma cells, non-pulmonary carcinomas, leukemias, and fibroblasts. A small-cell carcinoma cell line growing in suspension was found to have a different cytoskeletal composition from a variant cell line which grows attached to the culture dish. We have also identified differences in the cytoskeletal composition of adenocarcinomas of the lung compared to adenocarcinomas of other origins. The proteins characteristic of these different carcinomas are being isolated and purified by cutting protein spots from two-dimensional gels and elution into dialysis bags. The immunologic reactivity and peptide composition of these proteins will be compared with known cytoskeletal proteins. Monoclonal antibodies with specificity for nine cytoskeleton-associated proteins will be prepared. Two such monoclonal antibodies are currently being characterized. With these antibodies, the intracellular localization of these different proteins in cell lines, tumor tissues, and normal bronchial epithelium can be determined using immunofluorescence and immunoperoxidase-staining techniques. (E)